We have previously shown that supernatants derived from natural killer (NK) cells of HIV-seronegative individuals suppress HIV replication in human monocytes/macrophages (MO) and monocytic cell lines, but not in human T lymphocytes or T cell lines. Experiments have been undertaken to determine the nature of the factor(s) responsible for this suppression. Our results, thus far, show that the NK cell-derived supernatant contains Interferon- gamma (IFN-gamma) and granulocyte/monocyte colony stimulating factor (GM- CSF), and that each of these cytokines alone can suppress viral replication. Sequential removal of IFN-gamma (17) (kD) and GM-CSF (23 kD) from the supernatant using immunoaffinity gels did not eliminate the suppressive activity. In addition, size exclusion analysis revealed that the NK cell-derived suppressor factor has an apparent molecular weight greater than 30 kD. These data indicate that NK cells synthesize a greater than 30 kD HIV-suppressive factor distinct from IFN-gamma and GM-CSF. Work is in progress to identify and characterize this potentially novel factor.